Design Of Multi-targeting Self-assembling Peptides With Anticancer Activity And Study On Their Acting Mechanism

Cancer is the main cause of high morbidity and high mortality in the world.At present,surgery and chemotherapy are still the main treatments for cancer.The risk of surgery is high and the trauma to human body is serious,and it often leads to a series of complications.Chemical drugs may cause some toxic side effects to patients,and are prone to resulting in multiple drug resistance(MDR),which is not conducive to follow-up treatment.Therefore,it is imperative to develop a new class of anticancer drugs that do not have the toxicity of conventional chemotherapeutic drugs and are not affected by the common MDR mechanism.Compared with traditional chemotherapy drugs,peptides have obvious advantages.Anticancer peptides usually act on the cell membrane of tumor cells or induce apoptosis by disrupting mitochondrial membranes,thus being able to overcome MDR of tumor cells.However,anticancer peptides also have some shortcomings,such as short half-life in circulation,low stability and poor cell selectivity.Therefore,it is necessary to optimize the design of peptides in order to increase the application potential.In this study,the tumor-homing peptide RGDK was combined with the antitumor peptides,and the pro-drug strategy was used to incorporate specific protease-cleaving sites for the overexpressed proteases.An overall self-assembling tendency of the peptides was rationally designed to improve the targeting and stability of the antitumor peptides.According to the above design strategy,multi-targeting anticancer peptides(LIG series)were synthesized,including the tumor-targeting peptide,a linkage peptide segment and a functional peptide segment.Among them,the functional peptide is the CL-1 peptides(FLGALFRALSRLL)with antitumor activity,the target peptide is the RGDK sequence,and the cleavage site of matrix metalloproteinase IX(MMP-9)(PVGLIG)is integrated into the linking peptide.By probing 1,8-ANS fluorescence,three peptides(LIG-6,LIG-7 and LIG-8)were found to self-assemble in aqueous environment.They were able to form nanomicelles as observed by transmission electron microscopy.Their secondary structures in a hydrophobic environment were determined by circular dichroism(CD)asa-helix,while the secondary structures of LIG-6 and LIG-7 were random coil and that of LIG-8 was a-helix in an aqueous solution.LIG-8 had stronger activities on the MMP-9 overexpressed tumor cells,suggesting that the peptides may have be further activated by MMP-9.The serum stability of the self-assembling peptides was good.LIVE/DEAD cell staining experiments showed peptides LIG-6,LIG-7 and LIG-8 treat on HeLa cells by lysing cell membrane;The tumor-bearing nude mice model was established by subcutaneously injection of HeLa cells.After treatment with LIG-8 by para-tumor or intraperitoneal injection,a significant inhibitory effect on tumor growth was observed.The above experiments prove that the designed anti-tumor peptides with multiple targeting properties can effectively kill tumor cells,and have good serum stability.These multi-targeting self-assembling anticancer peptides may have promising clinical application potential.